This proposal investigates synaptic transmitters in mammalian taste buds. My Specific Aims include identifying the principal transmitters released by mouse taste cells during gustatory stimulation and determining where these transmitters act. Transmitters that have been identified to date and shown to be released by taste cells include serotonin, ATP, and norepinephrine. However, there is strong evidence that there are others, including glutamate, acetylcholine, and certain peptides. To complete the Aims, we will use a biosensor technique we developed and optimized in the previous funding period. Our biosensors consist of genetically-engineered CHO cells that are very sensitive to specific transmitters. We will use functional Ca2+ imaging to measure taste-evoked activity in taste cells and biosensors. We will correlate our findings with immunostaining for the receptors, transporters, and biosynthetic enzymes of candidate transmitters to confirm their presence and action in taste buds. This approach will allow us to answer such questions as: what are the transmitters in taste buds? Are there different transmitters for different tastes? How do taste cells interact via these transmitters? The long-term goal of this project is to construct a diagram of synaptic interactions that take place in the taste bud during taste stimulation. Understanding these interactions may help explain how different tastes (sweet, sour, salty, bitter, umami) are discriminated.